Increased reactivity of the paraventricular nucleus of the hypothalamus and decreased threat responding in male rats following psilocin administration.

Psychedelics have experienced renewed interest following positive clinical effects, however the neurobiological mechanisms underlying effects remain unclear. The paraventricular nucleus of the hypothalamus (PVN) plays an integral role in stress response, autonomic function, social behavior, and other affective processes. We investigated the effect of psilocin, the psychoactive metabolite of psilocybin, on PVN reactivity in Sprague Dawley rats. Psilocin increased stimulus-independent PVN activity as measured by c-Fos expression in male and female rats. Psilocin increased PVN reactivity to an aversive air-puff stimulus in males but not females. Reactivity was restored at 2- and 7-days post-injection with no group differences. Additionally, prior psilocin injection did not affect PVN reactivity following acute restraint stress. Experimental groups sub-classified by baseline threat responding indicate that increased male PVN reactivity is driven by active threat responders. These findings identify the PVN as a significant site of psychedelic drug action with implications for threat responding behavior.

Sex Differences in the Impact of Electronic Nicotine Vapor on Corticotropin-Releasing Factor Receptor 1 Neurons in the Mouse Ventral Tegmental Area.

Nicotine engages dopamine neurons in the ventral tegmental area (VTA) to encode reward and drive the development of nicotine addiction, however how nicotine alters a stress associated VTA population remains unclear. Here, we used male and female CRF1-GFP mice and nicotine vapor exposure to examine the effects of nicotine in VTA corticotropin-releasing factor receptor 1 (CRF1) neurons. We use immunohistochemistry and electrophysiology to examine neuronal activity, excitability, and inhibitory signaling. We found that VTA CRF1 neurons are mainly dopaminergic and project to the nucleus accumbens (NAc; VTA-NAcCRF1 neurons). VTA-NAcCRF1 neurons show greater phasic inhibition in naive females and greater focal nicotine-induced increases in firing in naive males. Following acute nicotine vapor exposure, phasic inhibition was not altered, but focal nicotine-induced tonic inhibition was enhanced in females and diminished in males. Acute nicotine vapor exposure did not affect firing in VTA-NAcCRF1 neurons, but females showed lower baseline firing and higher focal nicotine-induced firing. Activity (cFos) was increased in the CRF1 dopaminergic VTA population in both sexes, but with greater increases in females. Following chronic nicotine vapor exposure, both sexes displayed reduced basal phasic inhibition and the sex difference in tonic inhibition following acute vapor exposure was no longer observed. Additionally, activity of the CRF1 dopaminergic VTA population was no longer elevated in either sex. These findings reveal sex-dependent and exposure-dependent changes in mesolimbic VTA-NAc CRF1 neuronal activity, inhibitory signaling, and nicotine sensitivity following nicotine vapor exposure. These changes potentially contribute to nicotine-dependent behaviors and the intersection between stress, anxiety, and addiction.SIGNIFICANCE STATEMENT Nicotine is known to engage reward systems in the brain historically centering the neurotransmitter dopamine however, how nicotine impacts other neurons in the reward pathway is less clear. The current study investigates the impact of acute and chronic electronic nicotine vapor exposure in a genetically-defined cell population containing the stress receptor corticotropin-releasing factor 1 (CRF1) that is located in the reward circuitry. This study employs functional measures of neuronal activity and identifies important sex differences in nicotine's effects across time and exposure.

Chronic E-Cigarette Exposure Alters the Human Bronchial Epithelial Proteome.

RATIONALE: E-cigarettes vaporize propylene glycol/vegetable glycerin (PG/VG), nicotine, and flavorings. However, the long-term health effects of exposing lungs to vaped e-liquids are unknown. OBJECTIVES: To determine the effects of chronic vaping on pulmonary epithelia. METHODS: We performed research bronchoscopies on healthy nonsmokers, cigarette smokers, and e-cigarette users (vapers) and obtained bronchial brush biopsies and lavage samples from these subjects for proteomic investigation. We further employed in vitro and murine exposure models to support our human findings. MEASUREMENTS AND MAIN RESULTS: Visual inspection by bronchoscopy revealed that vaper airways appeared friable and erythematous. Epithelial cells from biopsy samples revealed approximately 300 proteins that were differentially expressed in smoker and vaper airways, with only 78 proteins being commonly altered in both groups and 113 uniquely altered in vapers. For example, CYP1B1 (cytochrome P450 family 1 subfamily B member 1), MUC5AC (mucin 5 AC), and MUC4 levels were increased in vapers. Aerosolized PG/VG alone significantly increased MUC5AC protein in human airway epithelial cultures and in murine nasal epithelia in vivo. We also found that e-liquids rapidly entered cells and that PG/VG reduced membrane fluidity and impaired protein diffusion. CONCLUSIONS: We conclude that chronic vaping exerts marked biological effects on the lung and that these effects may in part be mediated by the PG/VG base. These changes are likely not harmless and may have clinical implications for the development of chronic lung disease. Further studies will be required to determine the full extent of vaping on the lung.

Diacylglycerol lipase disinhibits VTA dopamine neurons during chronic nicotine exposure.

Chronic nicotine exposure (CNE) alters synaptic transmission in the ventral tegmental area (VTA) in a manner that enhances dopaminergic signaling and promotes nicotine use. The present experiments identify a correlation between enhanced production of the endogenous cannabinoid 2-arachidonoylglycerol (2-AG) and diminished release of the inhibitory neurotransmitter GABA in the VTA following CNE. To study the functional role of on-demand 2-AG signaling in GABAergic synapses, we used 1,2,3-triazole urea compounds to selectively inhibit 2-AG biosynthesis by diacylglycerol lipase (DAGL). The potency and selectivity of these inhibitors were established in rats in vitro (rat brain proteome), ex vivo (brain slices), and in vivo (intracerebroventricular administration) using activity-based protein profiling and targeted metabolomics analyses. Inhibition of DAGL (2-AG biosynthesis) rescues nicotine-induced VTA GABA signaling following CNE. Conversely, enhancement of 2-AG signaling in naïve rats by inhibiting 2-AG degradation recapitulates the loss of nicotine-induced GABA signaling evident following CNE. DAGL inhibition reduces nicotine self-administration without disrupting operant responding for a nondrug reinforcer or motor activity. Collectively, these findings provide a detailed characterization of selective inhibitors of rat brain DAGL and demonstrate that excessive 2-AG signaling contributes to a loss of inhibitory GABAergic constraint of VTA excitability following CNE.

VTA CRF neurons mediate the aversive effects of nicotine withdrawal and promote intake escalation.

Dopaminergic neurons in the ventral tegmental area (VTA) are well known for mediating the positive reinforcing effects of drugs of abuse. Here we identify in rodents and humans a population of VTA dopaminergic neurons expressing corticotropin-releasing factor (CRF). We provide further evidence in rodents that chronic nicotine exposure upregulates Crh mRNA (encoding CRF) in dopaminergic neurons of the posterior VTA, activates local CRF1 receptors and blocks nicotine-induced activation of transient GABAergic input to dopaminergic neurons. Local downregulation of Crh mRNA and specific pharmacological blockade of CRF1 receptors in the VTA reversed the effect of nicotine on GABAergic input to dopaminergic neurons, prevented the aversive effects of nicotine withdrawal and limited the escalation of nicotine intake. These results link the brain reward and stress systems in the same brain region to signaling of the negative motivational effects of nicotine withdrawal.

Nociceptin/orphanin FQ blockade of corticotropin-releasing factor-induced gamma-aminobutyric acid release in central amygdala is enhanced after chronic ethanol exposure.

BACKGROUND: The central nucleus of the amygdala (CeA) mediates stress- and addiction-related processes. Corticotropin-releasing factor (CRF) and nociceptin/orphanin FQ (nociceptin) regulate ethanol intake and anxiety-like behavior. In the rat, CRF and ethanol significantly augment CeA gamma-aminobutyric acid (GABA) release, whereas nociceptin diminishes it. METHODS: Using electrophysiologic techniques in an in vitro slice preparation, we investigated the interaction of nociceptin and CRF on evoked and spontaneous GABAergic transmission in CeA slices of naive and ethanol-dependent rats and the mechanistic role of protein kinase A. RESULTS: In neurons from naive animals, nociceptin dose-dependently diminished basal-evoked GABA(A) receptor-mediated inhibitory postsynaptic potentials (IPSPs) by decreasing GABA release and prevented, as well as reversed, CRF-induced augmentation of IPSPs, actions that required PKA signaling. In neurons from ethanol-dependent animals, nociceptin decreased basal GABAergic transmission and blocked the CRF-induced increase in GABA release to a greater extent than in naive controls. CONCLUSIONS: These data provide new evidence for an interaction between the nociceptin and CRF systems in the CeA. Nociceptin opposes CRF effects on CeA GABAergic transmission with sensitization of this effect in dependent animals. These properties of nociceptin may underlie its anti-alcohol and anxiolytic properties and identify the nociceptin receptor as a useful therapeutic target for alcoholism.